Phosphorylation of Lysosomal Enzymes in Fibroblasts. Marked Deficiency ofN-Acetylglucosamine-1-phosphotransferase in Fibroblasts of Patients with Mucolipidosis III

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Abstract
N-Acetylglucosamine-1-phosphotransferase activity was assayed in human skin fibroblasts using [.beta.-32P]UDP-N-acetylglucosamine as donor and dephosphorylated .beta.-N-acetyl-D-hexosaminidase as acceptor. An optimal transfer rate of N-acetylglucosamine 1-phosphate required CDP-choline and ADP to inhibit the breakdown of [.beta.-32P]UDP-N-acetylglucosamine, and a combination of leupeptin and iodoacetamide to protect the transferase. The transferase required Mg2+ or Mn2+. Using doubly labeled UDP-N-acetylglucosamine, simultaneous transfer of N-acetyl-[6-3H]glucosamine and [32P]phosphate to endogenous acceptors was demonstrated. Membranes prepared from fibroblasts from patients with mucolipidosis type III were defective in transfer of N-acetylglucosamine 1-phosphate. A residual transferase activity of < 10% of controls was detectable in fibroblast membranes of 8 patients with mucolipidosis type III. In membranes from fibroblasts from patients with mucolipidosis type II, N-acetylglucosamine-1-phosphotransferase activity was not detectable. The primary defect in mucolipidosis types II and III is apparently a deficiency in N-acetylglucosamine-1-phosphotransferase, the residual activity being higher in mucolipidosis type III than in mucolipidosis type II.

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